Effects of waterborne Pb/Cu mixture on Chinese toad, Bufo gargarizans tadpoles: morphological, histological, and intestinal microbiota assessment.

Coexistence of heavy metals in aquatic environments exert complex effects on amphibians. Here, the adverse effects of Pb (0.14 μM) combined with Cu at concentrations of 0, 0.25, and 1.0 μM were investigated in Bufo gargarizans tadpoles. Tadpoles were chronically exposed from Gosner stage (Gs) 26 to Gs 38, and morphology of tadpoles as well as intestinal histology and bacterial community were assessed. Our results indicated that Pb+Cu1.0 exposure induced significant retardation of somatic mass, total length, intestine mass, and intestine length as well as intestinal histological alterations. Pb+Cu0.25 and Pb+Cu1.0 exposure were associated with the loss of gut bacterial diversity. Proteobacteria and Bacteroidetes were two dominant phyla in tadpoles independently of heavy metal exposure, but the abundance of Proteobacteria increased significantly in Pb+Cu1.0 group and Bacteroidetes decreased significantly in all treatment groups. Furthermore, functional prediction indicated that metabolic disorders were associated with Pb+Cu0.25 and Pb+Cu1.0 exposure. Overall, relative limited shifts in intestinal bacterial diversity, composition, and functionality caused by Pb+Cu0 exposure, while coexistence of Pb and Cu induced gut dysbiosis and might further cause disturbance of metabolic homeostasis. The findings of this study provide insights into the effects of Pb and Cu coexistence on the health of amphibians.